The anaerobic oral bacterium, Bacteroides gingivalis, has been implicated as a major pathogen in adult periodontitis. Nutritional needs are of prime ecological importance in the localization of B. gingivalis to the gingival crevice area. B. gingivalis requires hemin for growth, however it is not known if other iron-compounds can fulfill this nutritional requirement. This study is designed to identify the mechanism of iron acquisition and utilization, and to assess their role in pathogenicity of B. gingivalis. The objectives of the proposed project are to define the iron requirements of B. gingivalis for growth in vitro and to relate these to growth in vivo. Iron sources will include iron bound to transferrin, lactoferrin, albumin, haptoglobin and hemopexin, sources commonly found in vivo. The mechanisms involved in iron acquisition will be examined by 1) determining if B. gingivalis can utilize siderophores produced by other organisms, 2) if B. gingivalis does indeed produce its own siderophores, and 3) if B. gingivalis produces specific proteins that are regulated by growth in iron- limited conditions. A genetic approach will be taken to examine more closely the requirement for iron, by constructing strains of B. gingivalis differing only in the ability to utilize hemin for growth. Tn4351 transpositional mutagenesis will be used to construct these strains. Isogenic strains will be used in our mouse model to assess the virulence of these strains. We will clone the corresponding genes for iron utilization using specific Tn4351-B. gingivalis mutant genomic DNA as probes in total genomic libraries. In addition, B. gingivalis grown in iron-sufficient and iron-deficient medium will be evaluated for its ability to cause infection in the mouse subcutaneous chamber model. These studies will allow for the evaluation of the role of iron utilization in the pathogenicity of B. gingivalis.